Bottom Line:
Treatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance.In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo.Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

ABSTRACTTreatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance. In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo. Suppression of SOX2 sensitizes cells to erlotinib-mediated apoptosis, ultimately decreasing the emergence of acquired resistance, whereas its ectopic expression reduces drug-induced cell death. We show that erlotinib relieves EGFR-dependent suppression of FOXO6, leading to its induction of SOX2, which in turn represses the pro-apoptotic BH3-only genes BIM and BMF. Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

Mentions:
Other transcripts induced or repressed following erlotinib treatment of mutantEGFR-addicted cells were not selective to EGFR signaling. Downregulated genesincluded known direct transcriptional targets of ERK signaling (CCND1, FOSL1,EGR1, IER3, IL-8) and shared feedback inhibitors of receptor tyrosinekinase (RTK) signaling (DUSP6) (Amit et al., 2007). This gene set overlaps with genes known to bedifferentially expressed following treatment of BRAF-mutant melanomacells with a MEK inhibitor and exposure of EGFR-mutant lung cancercells to an irreversible EGFR inhibitor (Figure1—figure supplement 3) (Kobayashiet al., 2006; Pratilas et al.,2009). In addition to SOX2, the 12 other transcripts induced by EGFRinhibition included genes encoding metabolizing enzymes (CYP1B1,CYP1A1) that are normally induced by treatment with a variety of smallchemical entities, genes that we also found to be equally well induced by inhibitionof downstream signaling pathways (MEK/MAPK inhibition using AZD6244 and PI3K/mTORinhibition using BEZ235), one transcript (CCNG2) previouslydescribed in another EGFR model (Kobayashi et al.,2006) and a long noncoding RNA (NEAT1). Taken alltogether, these results indicate that suppression of EGFR signaling in mutantEGFR-addicted lung cancer cells is highly specific in triggering transcriptionalinduction of SOX2.

Mentions:
Other transcripts induced or repressed following erlotinib treatment of mutantEGFR-addicted cells were not selective to EGFR signaling. Downregulated genesincluded known direct transcriptional targets of ERK signaling (CCND1, FOSL1,EGR1, IER3, IL-8) and shared feedback inhibitors of receptor tyrosinekinase (RTK) signaling (DUSP6) (Amit et al., 2007). This gene set overlaps with genes known to bedifferentially expressed following treatment of BRAF-mutant melanomacells with a MEK inhibitor and exposure of EGFR-mutant lung cancercells to an irreversible EGFR inhibitor (Figure1—figure supplement 3) (Kobayashiet al., 2006; Pratilas et al.,2009). In addition to SOX2, the 12 other transcripts induced by EGFRinhibition included genes encoding metabolizing enzymes (CYP1B1,CYP1A1) that are normally induced by treatment with a variety of smallchemical entities, genes that we also found to be equally well induced by inhibitionof downstream signaling pathways (MEK/MAPK inhibition using AZD6244 and PI3K/mTORinhibition using BEZ235), one transcript (CCNG2) previouslydescribed in another EGFR model (Kobayashi et al.,2006) and a long noncoding RNA (NEAT1). Taken alltogether, these results indicate that suppression of EGFR signaling in mutantEGFR-addicted lung cancer cells is highly specific in triggering transcriptionalinduction of SOX2.

Bottom Line:
Treatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance.In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo.Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

ABSTRACTTreatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance. In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo. Suppression of SOX2 sensitizes cells to erlotinib-mediated apoptosis, ultimately decreasing the emergence of acquired resistance, whereas its ectopic expression reduces drug-induced cell death. We show that erlotinib relieves EGFR-dependent suppression of FOXO6, leading to its induction of SOX2, which in turn represses the pro-apoptotic BH3-only genes BIM and BMF. Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.